Various scientific and scholarly articles are referenced in parentheses throughout the specification. These articles are incorporated by reference herein to describe the state of the art to which this invention pertains.
As the use of conventional pharmaceutical antibiotics (herein referred to as antibiotics) increases for medical, veterinary and agricultural purposes, the increasing emergence of antibiotic-resistant strains of pathogenic bacteria is an unwelcome consequence. This has become of major concern inasmuch as drug resistance of bacterial pathogens is presently the major cause of failure in the treatment of infectious diseases. Indeed, people now die of certain bacterial infections that previously could have been easily treated with existing antibiotics. Such infections include, for instance, Staphylococcus pneumoniae, causing meningitis; Enterobacter sp., causing pneumonia; Enterococcus sp., causing endocarditis, and Mycobacterium tuberculosis, causing tuberculosis.
The emergence of single- or multi-drug resistant bacteria results from a gene mobilization that responds quickly to the strong selective pressure that is a consequence of antibiotic uses. Over the last several decades, the increasingly frequent usage of antibiotics has acted in concert with spontaneous mutations arising in the bacterial gene pool to produce antibiotic resistance in certain strains. This gene pool is continually utilized by previously sensitive strains capable of accessing it by various means including the transfer of extrachromosomal elements (plasmids) by conjugation. As a result, single- and multi-drug resistance mutations are commonly found in a large variety of bacterial plasmids.
Presently there is no known method by which to avoid the selection of antibiotic resistant bacterial mutants that arise as a result of the many standard applications of antibiotics in the modern world. Accordingly, a need exists to develop alternative strategies of antibacterial treatment.
Interest in the use of bacteriophages to treat infectious bacterial diseases developed early in the twentieth century, and has undergone a resurgence in recent years. For instance, bacteriophages have been shown effective in the treatment of certain pathogenic E. coli species in laboratory and farm animals, and have been proposed as a viable alternative to the use of antibiotics (Smith & Huggins, J. Gen. Microbiol. 128: 307–318, 1981; Smith & Huggins, J. Gen. Microbiol. 129: 2659–2675, 1983; Smith et al., J. Gen. Microbiol. 133: 1111–1126, 1986; Kuvda et al., Appl. Env. Microbiol. 65: 3767–3773, 1999). However, the use of bacteriophages as antimicrobial agents has certain limitations. First, the relationship between a phage and its host bacterial cell is typically very specific, such that a broad host-range phage agent generally is unavailable. Second, the specificity of interaction usually arises at the point of the recognition and binding of phage to the host cell. This often occurs through the expression of surface receptors on the host cell to which a phage specifically binds. Inasmuch as such receptors are usually encoded by a single gene, mutations in the host bacterial cell to alter the surface receptor, thereby escaping detection by the phage, can occur with a frequency equivalent to or higher than, the mutation rate to acquire antibiotic resistance. As a result, if phage were utilized as commonly as antibiotics, resistance of pathogenic bacteria to phages could become as common a problem as antibiotic resistance.
Another approach to controlling pathogenic bacteria has been proposed, which relies on using molecular biological techniques to prevent the expression of antibiotic resistance genes in pathogenic bacteria (U.S. Pat. No. 5,976,864 to Altman et al.). In this method, a nucleic acid construct encoding an “external guide sequence” specific for the targeted antibiotic resistance gene is introduced into the pathogenic bacterial cells. The sequence is expressed, hybridizes with messenger RNA (mRNA) encoding the antibiotic resistance gene product, and renders such mRNA sensitive to cleavage by the enzyme RNAse P. Such a system also has limited utility, since it is targeted to specific antibiotic resistance genes. While the system may be effective in overcoming resistance based on expression of those specific genes, continued use of the antibiotics places selective pressure on the bacteria to mutate other genes and develop resistance to the antibiotic by another mechanism.
It is clear from the foregoing discussion that current alternatives to antibiotic use are limited and suffer many of the same drawbacks as antibiotic use itself. Thus, a need exists for a method of controlling pathogenic bacteria that is flexible in range and that cannot be overcome by the bacteria by a single small number of mutations.